During FY17 we accomplished the following: 1) Continued studies of mitochondrial function and autophagy in human CD4+ T cells to identify changes in these pathways during human aging. Change of autophagic flux during aging was monitored by the use of pharmacologic inhibitors that block the process at different stages. 2) Completed library preparation from 27 ATAC-Seq samples obtained from human monocytes from GESTALT. Six samples were sequenced to determine whether library quality was good enough to assay all 27 samples. Results were positive and the remaining 21 samples will be sent off for sequencing. 3) Continued to fractionate human PBMC into 11 cell types for transcriptomic, methylomic and proteomic analyses. With samples acquired over FY17, we are beyond the halfway mark in GESTALT. 4) Extended ATAC-Seq analysis to nave and memory B cell subsets in GESTALT. 5) Initiated studies of immune activation in GESTALT. First five donors (non-GESTALT) were used to standardize conditions to assay early (0-20 min) and late (18-20h) consequences of T and monocyte activation. For early time points, phospho-flow will be carried out by CyTOF and intracellular cytokine expression will be used to assay late activation events by multi-color flow cytometry. CyTOF will be carried out in collaboration with Matt Spitzer (UCSF) and multi-color flow will be carried out in collaboration with Pratip Chattopadhyay (NYU). Activation experiments were also carried out with total PBMC for comparison with purified cell types. 6) Initiated studies of autoimmune reactivity in rheumatoid arthritis patients using mass spectrometry with Dr. Gregory Ippolito (UT Austin).